The overall goal of this proposal is to define the control mechanisms that confer endometrial tissue specificity to the expression of the uteroferrin (UF) gene during pregnancy in the pig. The molecular basis for the interaction of cis-acting elements located 5' of the UF gene with nuclear transacting factors to achieve high levels of transcription in the uterus will be analyzed to provide further understanding of the mechanisms involved in the appropriate expression of endometrial genes, whose products are essential for embryonic, fetal and placental development, uterine growth and tissue remodelling. The endometrial-specific response element (ESRE), which represents the sequences conferring endometrial-specific expression of the UF gene, will be identified by deletion mapping and linker-scanning mutagenesis of the UF gene 5' flanking region and analysis of the corresponding mutated fragments for UF promoter activity. Studies will utilize a transient expression system in the hormone-responsive endometrial epithelial cell line (HRE-H9), which supports UF gene promoter activity. The transacting factor (binding protein) that binds to ESRE will be characterized in uterine endometrium and in non-UF expressing tissues of pregnant gilts to define the specificity of their binding interactions. The ESRE will be used as probe to isolate the complementary DNA (cDNA) corresponding to the transacting factor from a lambda gt11 porcine endometrial expression library. This clone will be characterized by standard molecular biology techniques to deduce the amino acid sequence of the ESRE-binding protein and used as probe in Northern blot analysis to evaluate the pattern of expression of the mRNA encoding this protein relative to that of UF, during uterine and fetal development and as a function of hormonal status in gilts. Finally, the cDNA for ESRE-binding protein and the UF-CAT chimeric gene will be co- expressed in HRE-H9 cells and in non-UF expressing cells (i.e. AKR-2B and JEG-3) to demonstrate the functional ability of the transacting factor in the endometrial-specific expression of the UF gene. Uteroferrin gene is an excellent candidate gene to elucidate the mechanisms involved in the control of uterine gene expression. Results from these studies will clarify essential mechanisms underlying maternal-fetal interactions during critical periods of conceptus and fetal development.